The present invention is directed to solubilizing compounds, a device and a method for solubilizing and removing carboxylic acids and especially fatty acids from oils, fats, aqueous emulsions, aqueous or organic solutions. Devices utilizing the inventive method shall be used for separating carboxylic acids from oils, fats, aqueous emulsions, lipophilic media, aqueous media or organic solutions, respectively, thus changing their reaction conditions. One application is a device for removing fatty acids from the blood of subjects in the need thereof. It shall be used further for analytic, respectively diagnostic purposes of fatty acid concentrations in corporal fluids of subjects, foods or pharmaceutical preparations. Furthermore, this technique shall be applied for the removal of carboxylic acid residues in industrial solutions, for example as arising in the food and oil industry.
In general, fatty acids are highly lipophilic molecules which are barely soluble in aqueous solutions. Therefore only small concentrations of fatty acids can be solubilized in aqueous solutions while all fatty acid molecules exceeding this concentration are present in form of micelles, form an emulsion by phase separation, or are absorbed to container walls and/or other lipophilic or amphilic molecules such as proteins in the solution. Above the critical micelle concentration (CMC) of esterified and non-esterified carboxylic acids the concentration of free fatty acids in an aqueous medium remains unchanged.
Fatty acids tend to form emulsions in aqueous media. In presence of proteins or cellular structures fatty acids can be absorbed by them or adsorb to them. Solubilizing of such immobilized fatty acids mainly depends on the critical micelle concentration (CMC) of the fatty acid in the surrounding aqueous medium. Emulators and detergents are able to increase the CMC of hydrophobic substances and thus help to detach immobilized lipophilic molecules. These emulators and detergents may convert emulsions into mini-, micro- or nano-emulsions. Herein the contacting area of the solubilised fatty acids with the aqueous phase is increased. This allows for a better separability and extractability of the solubilised fatty acids. Thus also the reactivity with other molecules is augmented.
Emulsions of esterified and non-esterified fatty acids with an aqueous medium can be completely separated only by means of an organic solvent. Without the help of a membrane this can only be achieved by transferring the fatty acids into an organic phase by mixing with an organic solvent. Extraction is also possible by adsorption to an acceptor. In the presence of adsorber molecules such as proteins the separation of fatty acids in an emulsion or suspension by phase separation or extraction often is incomplete. Furthermore, the capacity of this technique is limited and usually not suited for online (continuous) processing. When filtering such emulsions the aqueous fraction can be filtered almost entirely. However, also hydrophilic molecules, in particular large proteins, are retained and separated along with the organic phase. A molecular separation can be achieved with chromatographic methods. These methods, however, are time-consuming and limited in their capacity.
Another method to separate carboxylic acids from aqueous or organic media is distillation. However, this procedure has a high energy demand and might generate isomerization of the carboxylic acids or denaturate organic components within the medium. A further method is saponification. Added salts are often difficult to remove from the organic as well as from the aqueous solution during further processing.
Thus there is a need for a continuous and selective extraction of fatty acids from emulsions of aqueous or organic solutions. The aim of the present invention is to provide a simple, quick and biocompatible separation of fatty acids from aqueous emulsions or organic media.
Surprisingly, it was found that this aim can be achieved by adding a solubilizing compound to the aqueous emulsion or aqueous media such as blood, lipophilic medium or organic medium containing the carboxylic acids or mixtures of carboxylic acids with other organophilic molecules. An inventive solubilizing compound having the characteristics as defined herein is able to solubilize carboxylic acids and convert the emulgated carboxylic acids into micro- or nanoemulsions which allow a separation by means of separation methods such as dialysis, filtration and electrophoresis.
Thus the task is solved by the ensuing technical teachings of the independent claims of the present invention. Further advantageous embodiments of the invention result from the dependent claims, the description and the examples.
Fatty Acids
In general, fatty acids have a carboxylic head group and a long aliphatic chain. Depending on the presence of double bonds they are differentiated into saturated and unsaturated fatty acids. There are differing definitions in literature about fatty acids. One definition states that carboxylic acids with 4 carbon atoms or higher are regarded as fatty acids. Naturally occurring fatty acids, however, have at least 8 carbon atoms. At these carbon atoms at least one nitro group can replace hydrogen atom(s) and turn them into nitro-fatty acids. Also nitro-fatty-acids may carry further substituents, such as listed above.
Examples for linear saturated fatty acids are octanoic acid (caprylic acid), decanoic acid (caprinic acid), dodecanoic acid (lauric acid), tetradecanoic acid (myristic acid), hexadecaoic acid (palmitic acid), heptadecanoic acid (margaric acid), octadecanoic acid (stearic acid), eicosanoic acid (arachidic acid), docosanoic acid (behenic acid) and tetracosanoic acid (lignoceric acid).
According to the invention a preferred subgroup of the saturated fatty acids to be separated are myristic acid, palmitic acid, and stearic acid.
Examples for monoolefinic fatty acids are cis-9-tetradecenoic acid (myristoleic acid), cis-9-hexadecenoic acid (palmitoleic acid), cis-6-hexadecenoic acid (salpenic acid), cis-6-octadecenoic acid (petroselinic acid), cis-9-octadecenoic acid (oleic acid), cis-11-octadecenoic acid (vaccenic acid), 12-hydroxy-9-cis-octadecenoic acid (ricinoleic acid), cis-9-eicosenoic acid (gadoleinic acid), cis-1′-eicosenoic acid (gondoic acid), cis-13-docosenoic acid (erucic acid), cis-15-tetracosenoic acid (nervonic acid), t9-octadecenoic acid (elaidic acid), t11-octadecenoic acid (t-vaccenic acid) and t3-hexadecenoic acid. According to the invention a preferred subgroup of the unsaturated fatty acids to be separated are the trans-isomers t9-octadecenoic acid, t11-octadecenoic acid, and t3-hexadecenoic acid.
Examples for polyolefinic fatty acids are 9,12-octadecadienoic acid (linoleic acid), 6,9,12-octadecatrienoic acid (γ-linoleic acid), 8,11,14-eicosatrienoic acid (dihomo-γ-linoleic acid), 5,8,11,14-eicosatrienoic acid (arachidonic acid), 7,10,13,16-docosatetraenoic acid, 4,7,10,13,16-docosapentaenoic acid, 9,12,15-octadecatrienoic acid (α-linolenic acid), 6,9,12,15-octadecatetraenic acid (stearidonic acid), 8,11,14,17-eicosatetraenoic acid, 5,8,11,14,17-eicosapentaenoic acid (EPA), 7,10,13,16,19-docosapentaenoic acid (DPA), 4,7,10,13,16,19-docosahexaenic acid (DHA), 5,8,11-eicosatrienoic acid (mead acid), 9c 11t 13t eleostearinoic acid, 8t 10t 12c calendic acid, 9c 11t 13c catalpic acid, 4, 7, 9, 11, 13, 16, 19 docosaheptadecanoic acid (stellaheptaenoic acid), taxolic acid, pinolenic acid and sciadonic acid.
According to the invention a preferred subgroup of the unsaturated fatty acids to be separated are the trans-isomers of linoleic acid, γ-linoleic acid, EPA, and DPA.
Examples for acetylenic fatty acids are 6-octadecinoic acid (tariric acid), t11-octadecen-9-ynoic acid (santalbic or ximenic acid), 9-octadecynoic acid (stearolic acid), 6-octadecen-9-ynoic acid (6,9-octadecenynoic acid), t10-heptadecen-8-ynoic acid (pyrulic acid), 9-octadecen-12-ynoic acid (crepenic acid), t7,t11-octadecadiene-9-ynoic acid (heisteric acid), t8,t10-octadecadiene-12-ynoic acid, 5,8,11,14-eicosatetraynoic acid (ETYA).
It shall be noted that according to the invention also the bases, respectively salts of the aforementioned fatty acids shall be subsumed under the general terms fatty acids or free fatty acids.
Examples for suitable organic and inorganic bases for salt formation are bases derived from metal ions, e.g., aluminum, alkali metal ions, such as sodium of potassium, alkaline earth metal ions such as calcium or magnesium, or an amine salt ion or alkali- or alkaline-earth hydroxides, -carbonates or -bicarbonates. Examples include aqueous sodium hydroxide, lithium hydroxide, potassium carbonate, ammonia and sodium bicarbonate, ammonium salts, primary, secondary and tertiary amines, such as, e.g., lower alkylamines such as methylamine, t-butylamine, procaine, ethanolamine, arylalkylamines such as dibenzylamine and N,N-dibenzylethylenediamine, lower alkylpiperidines such as N-ethylpiperidine, cycloalkylamines such as cyclohexylamine or dicyclohexylamine, morpholine, glucamine, N-methyl- and N,N-dimethylglucamine, 1-adamantylamine, benzathine, or salts derived from amino acids like lysine, ornithine or amides of originally neutral or acidic amino acids or the like.
The following carboxylic acids are preferred examples of fatty acids:
octanoic acid (caprylic acid), decanoic acid (capric acid), dodecanoic acid (lauric acid), tetradecanoic acid (myristic acid), hexadecanoic acid (palmitic acid), heptadecanoic acid (margaric acid), octadecanoic acid (stearic acid), eicosanoic acid (arachidic acid), docosanoic acid (behenic acid), tetracosanoic acid (lignoceric acid), cis-9-tetradecenoic acid (myristoleic acid), cis-9-hexadecenoic acid (palmitoleic acid), cis-6-octadecenoic acid (petroselinic acid), cis-9-octadecenoic acid (oleic acid), cis-11-octadecenoic acid (vaccenic acid), cis-9-eicosenoic acid (gadoleic acid), cis-11-eicosenoic acid (gondoic acid), cis-13-docosenoic acid (erucic acid), cis-15-tetracosenoic acid (nervonic acid), t9-octadecenoic acid (elaidic acid), t11-octadecenoic acid (t-vaccenic acid), t3-hexadecenoic acid, 9,12-octadecadienoic acid (linoleic acid), 6,9,12-octadecatrienoic acid (γ-linoleic acid), 8,11,14-eicosatrienoic acid (dihomo-γ-linolenic acid), 5,8,11,14-eicosatetraenoic acid (arachidonic acid), 7,10,13,16-docosatetraenoic acid, 4,7,10,13,16-docosapentaenoic acid, 9,12,15-octadecatrienoic acid (α-linoleic acid), 6,9,12,15-octadecatetraenoic acid (stearidonic acid), 8,11,14,17-eicosatetraenoic acid, 5,8,11,14,17-eicosapentaenoic acid (EPA), 7,10,13,16,19-docosapentaenoic acid (DPA), 4,7,10,13,16,19-docosahexaenoic acid (DHA), 5,8,11-eicosatrienoic acid (mead acid), 9c 11t 13t eleostearic acid, 8t 10t 12c calendic acid, 9c 11t 13c catalpic acid, 4, 7, 9, 11, 13, 16, 19 docosaheptadecanoic acid (stellaheptaenoic acid), taxoleic acid, pinolenic acid, sciadonic acid, 6-octadecynoic acid (tariric acid), t1′-octadecen-9-ynoic acid (santalbic or ximenynic acid), 9-octadecynoic acid (stearolic acid), 6-octadecen-9-ynoic acid (6,9-octadecenynoic acid), t10-heptadecen-8-ynoic acid (pyrulic acid), 9-octadecen-12-ynoic acid (crepenynic acid), t7,t11-octadecadiene-9-ynoic acid (heisteric acid), t8,t10-octadecadiene-12-ynoic acid, 5,8,11,14-eicosatetraynoic acid (ETYA), eleostearic acid, calendic acid, catalpic acid, stellaheptaenoic acid, taxoleic acid, retinoic acid, isopalmitic acid, pristanic acid, phytanic acid, 11,12-methyleneoctadecanoic acid, 9,10-methylenhexadecanoic acid, coronaric acid, (R,S)-lipoic acid, (S)-lipoic acid, (R)-lipoic acid, 6,8-bis(methylsulfanyl)-octanoic acid, 4,6-bis(methylsulfanyl)-hexanoic acid, 2,4-bis(methylsulfanyl)-butanoic acid, 1,2-dithiolane carboxylic acid, (R,S)-6,8-dithiane octanoic acid, (R)-6,8-dithiane octanoic acid, (S)-6,8-dithiane octanoic acid, cerebronic acid, hydroxynervonic acid, ricinoleic acid, lesquerolic acid, brassylic acid and thapsic acid.
Fatty Acids in Blood
In mammals fatty acids serve as physiologically important energy substrates and play a critical role in energy metabolism. Moreover, they are important substrates for the synthesis of membrane phospholipids and biologically active agents like eicosanoids and leukotrienes. The mammalian body heavily relies on fatty acids as suppliers of chemically stored energy, building blocks of cellular membranes and signal transducers. The main source of fatty acids is dietary lipid, digested in the gastro-intestinal tract by the catalytic action of pancreatic hydrolytic enzymes. Part of fatty acids is produced by the liver taking carbohydrates as substrate. A large percentage of fatty acids, however, is stored in fat cells (adipocytes) composing adipose tissue in form of triacylglycerol.
The concentration of esterified and unesterified fatty acids in the blood depends on several factors such as food intake or release from adipose tissue. Fatty acids can be bound or attached to other molecules, such as in triglycerides or phospholipids, or to a smaller percentage fatty acids occur unbound. In any case, fatty acids are insoluble in water and must be bound to a water soluble component for transport in the organism. Fatty acids are transported in the body via the lymphatic and vascular system. Basically, two transport forms are at hand: Fatty acids can be transported as triacylglycerols, which is the main component of circulating lipoproteins such as chylomicrons and very-low density lipoproteins, or as non-esterified fatty acids that are bound to plasma proteins, in particular plasma albumin. Free fatty acids that are completely unbound have a very low solubility and only occur in very low concentrations.
The composition, distribution and concentration of fatty acids in human blood can vary a lot and is made up by the sum of the different fractions of plasma: Cholesterol ester, phospholipids, and triacylglycerols as well as albumin-bound fatty acids. The saturated fatty acids in human blood are mostly made up by myristic acid (14:0), palmitic acid (16:0) and stearic acid (18:0). The main type of monounsaturated fatty acids belong to the group of oleic acid (18:1) and palmitoleic acid (16:1). Polyunsaturated omega-3 fatty acids include linolenic acid (18:3), eicosapentaenoic acid (20:5), docosapentaenoic acid (22:5) and docosahexaenoic acid (22:6). Polyunsaturated omega-6 fatty acids are mostly linoleic acid (18:2), eicosadienoic acid (20:2), dihomogammalinolenic acid (20:3), arachidonic acid (20:4), adrenic acid (22:4) and docosapentaenoic acid (22:5). The concentration of other fatty acids is usually very low in whole blood, but can vary depending on genetics, nutrition and life style.
Blood fatty acids concentrations are increased in obese patients and contribute to type 2 diabetes, hepatic steatosis and several cardiovascular disorders such as atherosclerosis. The pathogenetic role of fatty acids in the development of athrosclerosis and associated diseases such as cerebral, myocardal, renal, erectile dysfunction has been elucidated. Not intended to be comprehensive, some aspects should be outlined in the following. An elevation of fatty acids was found to be responsible for increase of reactive oxygen radical formation causing endothelial dysfunction which can be attenuated by an antioxidant (Pleiner et al, FFA-induced endothelial dysfunction can be corrected by vitamin C. J Clin Endocrinol Metab 2002, 87, 2913-7). This effect is increased by trans-fatty acids which are suspected to have additional deliterious effects (Lopez Garcia et al, Consumption of trans-fatty acids is related to plasma biomarkers of inflammation and endothelial dysfunction. J Nutr 2005, 135, 562-566; Mozaffarian et al, Health effects of trans-fatty acids: experimental and observational evidence. Eur J Clin Nutr 2009, 63 Suppl 2, S5-21). They are accused to increase blood pressure and found to be a pathogenetic factor in arterial hypertension (Zheng et al, Plasma fatty acid composition and 6-year incidence of hypertension in middle-aged adults: the Atherosclerosis Risk in Communities (ARIC) Study. Am J Epidemiol 1999, 150, 492-500). Trans-fatty acids were found to increase the risk of myocardial infarction and sudden heart death (Ascherio et al, Trans-fatty acids intake and risk of myocardial infarction. Circulation 1994, 89, 94-101; Baylin et al, High 18:2 trans-fatty acids in adipose tissue are associated with increased risk of nonfatal acute myocardial infarction in Costa Rican adults. J Nutr 2003, 133, 1186-1191). Together with a chronic elevation of fatty acid blood concentrations they are responsible for insulin resistance and development of diabetes mellitus (Krachler et al, Fatty acid profile of the erythrocyte membrane preceding development of Type 2 diabetes mellitus. Nutr Metab Cardiovasc Dis 2008, 18, 503-510; Lionetti et al, From chronic overnutrition to insulin resistance: the role of fat-storing capacity and inflammation. Nutr Metab Cardiovasc Dis 2009, 19, 146-152; Yu et al, Mechanism by which fatty acids inhibit insulin activation of insulin receptor substrate-1 (IRS-1)-associated phosphatidylinositol 3-kinase activity in muscle. J Biol Chem 2002, 277, 50230-50236). Altogether an increased turn-over of fatty acids as a result of chronic overnutrition is now believed to be the most important pathomechanism in the development of the most common diseases in industrialized countries (Bays, “Sick fat,” metabolic disease, and atherosclerosis. Am J Med 2009, 122, S26-37). Medical treatment for effective reduction of overweight is lacking (Aronne et al, When prevention fails: obesity treatment strategies. Am J Med 2009, 122, S24-32). However, obese persons who succeed in reducing body weight and thus a significant reduction of fatty acid-induced disorders can be found (Lien et al, The STEDMAN project: biophysical, biochemical and metabolic effects of a behavioral weight loss intervention during weight loss, maintenance, and regain. Omics 2009, 13, 21-35; Schenk et al, Improved insulin sensitivity after weight loss and exercise training is mediated by a reduction in plasma fatty acid mobilization, not enhanced oxidative capacity. J Physiol 2009, 587, 4949-4961). Therefore a medical device to effectively reduce the total amount of fatty acids and preferably those with increased pathogenicity is desirable.
Surgical extraction of subcutaneous adipose tissue was found to be ineffective in reducing circulating fatty acid concentrations or their qualitative content. Removal of the lipoprotein fraction carrying high concentrations of cholesterol by direct adsorption from blood can be accomplished by adsorption or filtration of these particles. Those procedures for online blood purification are called LDL apheresis. While designed to reduce LDL cholesterol, they also adsorb triglycerides. However, the quantity of triglycerides extracted is not sufficient for an effective reduction of the body content of fatty acids.
The fatty acid content of blood is low in the fastening state at rest. However, significant rise is observed during lipolysis (see below). Due to the insolubility in an aqueous medium the transport of unesterified fatty acids is accomplished by proteins and cellular structures (Spector et al, Utilization of long-chain free fatty acids by human platelets. J Clin Invest 1970, 49, 1489-1496). The major transport protein in blood is albumin. The presence of at least 10 specific binding sites for fatty acids has been documented. However, the binding capacity might increase dramatically by formation of micellar structures with fatty acids in a condition of excess of fatty acids or other lipids (Schubiger et al, Mixed micelles: a new problem-free solution for omega-123I-heptadecanoic acid in comparison. Nuklearmedizin 1984, 23, 27-28).
With a molarity of albumin of about 600 μmol/l a binding capacity of at least 0.006 mol/l for fatty acids would exist which equals about 0.0035 kg/l (Berk and Stump, Mechanisms of cellular uptake of long chain free fatty acids. Mol Cell Biochem 1999, 192, 17-31).
Furthermore, fatty acids are transported in esterified form as mono-, di- or triacyl glycerols. The fastening serum concentration varies considerably. However, normal values are set to be below 150 mg/dl (1.7 mmol/l). Postprandially or during exercising the concentration can rise several-fold and even exceed 1000 mg/dl (11.3 mmol/l).
Only spare reports exist which investigate the differences in the lipid content at various sites within the circulation. In these investigations significant higher values for fatty acids and triglycerides were found to be present in the central venous system (Vena cava) as compared to other measuring sites (Wiese et al, Lipid composition of the vascular system during infancy, childhood, and young adulthood, J. Lipid Res. 1967, 8, 312-320; Zauner et al, Pulmonary arterial-venous differences in lipids and lipid metabolites. Respiration 1985, 47, 214-219). No reports exist about the changes of the central venous lipid content during exercise and induced lipolysis.
It was found now, that during physical excercising the lipid content sharply increases in the central abdominal veins exhibiting an increasing difference in the lipid content between the central as compared to a peripheral assess site as described below.
Thus reducing the fatty acid content in blood using the methods and the devices and the solubilizing compounds disclosed herein is useful to treat the diseases mentioned above associated with an elevated level of fatty acids in the blood or in the organism.
Thus the present invention relates to the treatment and prophylaxis of fatty acid-induced disorders such as type 2 diabetes, hepatic steatosis, cardiovascular disorders such as arterial hypertension, myocardial infarction, stroke, sudden heart death, atherosclerosis, diseases associated with atherosclerosis such as cerebral, myocardial, renal and erectile dysfunction, as well as to weight reduction and cholesterol reduction and also to the prevention of insulin resistance and the prevention of the development of diabetes mellitus by using the solubilizing compounds disclosed herein in order to remove fatty acids from the blood.
Lipolysis
Plasma fatty acids are an important energy substrate. The availability of fatty acids is determined predominantly by their mobilization from adipose tissue triacylglycerol stores by the process of lipolysis. In man, lipolysis of adipose tissue is regulated by a number of hormonal, paracrine and/or autocrine signals. The main hormonal signals may be represented by catecholamines, insulin, growth hormone, natriuretic peptides, thyroxine, and some adipocytokines (Stich and Berlan, Physiological regulation of NEFA availability: lipolysis pathway. Proc Nutr Soc 2004, 63, 369-374). The absolute levels and relative importance and contribution of these signals vary in different physiological situations, with diet and physical exercise being the main physiological variables that affect hormonal signalling. A family of enzymes called lipases with distinct functions is responsible for the breakdown of triglycerides stored within fat cells for energy storage. Carbonhydrates and fatty acids are the major energy fuels for muscle contraction. During exercise training lipolysis liberates 7.1+/−1.2 micromol×min(−1)×kg(−1) body weight, which would result in a release of fatty acids of 4200 μmol per hour in a person with a weight of 100 kg which equals 0.15 kg fatty acids (Coggan et al, Fat metabolism during high-intensity exercise in endurance-trained and untrained men. Metabolism 2000, 49, 122-128). However, stimulation of lipolysis by pharmacological intervention and/or local physical measures may further increase lipolytic capacity. Lipolysis was increased up to 3-fold by systemic application of natural receptor agonists or drugs (Riis et al, Elevated regional lipolysis in hyperthyroidism. J Clin Endocrinol Metab 2002, 87, 4747-4753; Barbe et al, In situ assessment of the role of the beta 1-, beta 2- and beta 3-adrenoceptors in the control of lipolysis and nutritive blood flow in human subcutaneous adipose tissue. Br J Pharmacol 1996, 117, 907-913). Adrenoreceptor agonists exhibiting stimulating lipolysis are: Adrenaline, noradrenaline, isoprenaline, ephedrine, isoproteriol, salbutamol, theophylline, fenoterol, orciprenaline, a.o.
Lipolytic effects have also been described from physical alterations of fat tissue. Researchers found that ultrasound had a liquidifying effect on adipose tissue leading to a reduced content of fat tissue when performed during starvation (Faga et al, Ultrasound-assisted lipolysis of the omentum in dwarf pigs. Aesthetic Plast Surg 2002, 26, 193-196; Miwa et al, Effect of ultrasound application on fat mobilization. Pathophysiology 2002, 9, 13).
Though for all measures mentioned above increase in lipolysis has been documented the measurable effect on concentrations of unesterified fatty acids was small. In a pilot investigation it was found that after stimulation of lipolysis the content of fatty acids tremendously increased when measured by using the intentive method for solubilisation of fatty acids. Furthermore, it was found that the content of fatty acids was much higher within the abdominal venous system than in the peripheral circulation. This finding is surprising, since it has not been observed in animal studies when measuring various blood collections sites simultaneously.
Therefore, stimulation of lipolysis while performing blood purification from fatty acids by the intentive procedure and using an abdominal central vene as an access site is a preferred embodiment of the invention.
The extraction fraction could be increased if the content of esterified and unesterified fatty acids transported in blood could be elevated while performing the procedure.
Surprisingly, this task could be solved by increasing lipolysis through the inventive method.
Solvation and Adhesion Behavior of Fatty Acids in Aqueous Media
The solubility of carboxylic acids in water is minimal when the length of the carbon chain exceeds 4 carbon atoms and in absence of hydroxy (—OH) groups, carboxyl (—COOH) groups or other hydrophilic polar or charged groups and/or by introduction of alkyl substituents or other lipophilic groups.
Solubility can be increased by detergents which penetrate fatty acid micelles thereby reducing their stability and reducing their size, and increasing the number of free fatty acid molecules in the aqueous medium. Both free fatty acids and micelles tend to bind to lipotropic structures. Among those are carbon, metals, ceramics, natural and synthetic polymers. Furthermore, organic structures carry lipophilic regions, some of which are designated to specifically bind fatty acids, which form membrane or lipid transport proteins. The steric binding sites are mostly lined by hydrophobic amino acids.
In blood, lipids are electrostatically bound to specialized transport proteins. Fatty acids are mainly transported by albumin. The binding of fatty acids at the albumin molecule relies also on electrostatic forces which are localized in hydrophobic pockets. The binding energy of those pockets varies, however the pKa for all of them is substantially higher than the CMC of the fatty acids. Therefore, fatty acids remain in the surrounding medium even after complete removal of free fatty acids. Extraction of fatty acids from albumin was found to be almost complete when organic solvents were used for their liberation because of the better dissolution in organic solvents. However, those solvents alter the protein structure making them unsuitable for further processing or use in a living organism. To use albumin for medical or other purposes it is necessary to reduce their fatty acid content without altering the structure and functionality of albumin. This task can be solved by activated carbon particles which possess a higher binding affinity to fatty acids than albumin. However, this process needs further steps for purification of albumin. Therefore up to now there is no procedure that allows quick liberation and solubilization of the whole fatty acid content of an albumin molecule within an aqueous medium which does not alter the ultrastructure and function of the albumin molecules.
Carboxylic acids are also transported within phospholipid vesicles. Electrostatic interactions between the hydrocarbon chains of the carboxylic acids and those of the phospholipids retain carboxyl acids from diffusion to a surrounding aqueous medium. Mutatis mutandis, this applies also for other organic solutions, biomasses or organic waste waters. In organic solutions destinated for further refinement, purification or use where it is desirable not to use an organic solvent, an alternative biocompatible procedure is desirable. So far such a procedure is lacking.
Surprisingly, this aim can be reached by the use of at least one solubilizing compound as disclosed herein comprising at least one amidino and/or at least one guanidino moiety and especially solubilizing compounds of general formula (I), (II) and (III) and most especially arginine and derivatives thereof.
The carboxylic acids which shall be removed are normally contained in an aqueous medium or aqueous solution such as blood or blood plasma or in an aqueous emulsion such as milk or in an organic medium such as fuel, gas, bio-diesel, gasoline, petrol and the like or in oils such as vegetable oils like linseed oil, walnut oil, flax oil, evening primrose oil, sunflower oil, sunflower seed oil, soybean oil, rapeseed oil, olive oil, virgin olive oil, palm oil, palm kernel oil, peanut oil, cottonseed, coconut oil, corn oil, grape seed oil, hazelnut oil, rice bran oil, safflower oil, sesame oil as well as animal oils such as fish oil or contained in fats such as butter, oleo or margarine.
In case the carboxylic acid is contained in water, an aqueous medium, an aqueous emulsion or an aqueous suspension, the at least one solubilizing compound can be directly added to the aqueous medium, emulsion or suspension or the at least one solubilizing compound can be dissolved in water and this aqueous solution can be added to the aqueous medium, emulsion or suspension containing the carboxylic acids. After this addition the formation of a nanoemulsion and/or microemulsion is observed.
In case the carboxylic acids are contained in an organic medium or a lipophilic organic medium, the solubilizing compound is dissolved in water and the solution of the solubilizing compound in water is added to the organic medium. A two phase mixture is obtained and the carboxylic acids are transferred into the aqueous phase. It is assumed that a complex or aggregate of one molecule carboxylic acid with one molecule solubilizing compound or a dimer or trimer thereof is formed which makes the carboxylic acid soluable in water. Thus it is preferred to stir or shake the two phase mixture of the organic and aqueous layer in order to obtain an intensive mixing of the two layers. The carboxylic acids contained in the aqueous phase can be removed by phase separation. If desired, the extraction method can be repeated.
In case the carboxylic acids are contained in an oil or fat, the solubilizing compound is dissolved in water and the solution of the solubilizing compound in water is added to the oil or fat. If desired, an organic solvent could be added to the oil or fat in order to reduce viscosity of the oil or fat to make the oil or fat better stirrable. The mixture of the oil or fat and the aqueous solution of the solubilizing compound is stirred. The carboxylic acid is transferred into the aqueous phase and the aqueous phase can be removed by decantation or phase separation. The extraction process can be repeated for several times if desired.
Thus the invention also relates to an aqueous microemulsion and/or an aqueous nanoemulsion containing at least one solubilizing compound and at least one carboxylic acid in a microemulgated or in a nanoemulgated form.
If the solubilizing compound is used in an excess of 1.2 to 2.8, preferably 1.5 to 2.5 and more preferably in an excess of 1.7 to 2.3 mol equivalents, it is possible to remove more than 90% of the carboxylic acids in one extraction step. If the extraction step is repeated twice, up to 99% of the carboxylic acids can be removed.
The carboxylic acids which can be removed are especially carboxylic acids with more than 5 carbon atoms, more preferably with more than 7 carbon atoms and especially preferred with more than 9 carbon atoms. Preferably the carboxylic acids are fatty acids as disclosed herein while also other lipophilic compounds containing a carboxyl group or carboxylic acid group such as drugs or toxines can be removed by this method. One carboxylic acid which is explicitly disclaimed from the present invention is naproxen. Moreover it is not the intention of the present invention to provide methods and compounds or devices for the solubilization of pharmaceuticals in order to prepare galenic formulations. Especially preferred is the removal and solubilization of naphthenic acid from oil, petrol, gas and fuel. Moreover preferred carboxylic acids are such carboxylic acids which contain double and/or triple bonds such as unsaturated and polyunsaturated fatty acids. Still more preferred are physiologic carboxylic acids and especially these physiologic carboxylic acids which occur in human beings. For industrial purposes the unsaturated fatty acids are preferably removed and solubilized from the source material such as oils and fats while for medical purposes the saturated fatty acids are preferably removed from the blood of the patient. Moreover these carboxylic acids are preferred which occur in oil and fats of the above mentioned origin, especially from animals such as fishes, corn, olives, corn, crop, rice, soya and the like. In case the carboxylic acids which shall be removed from the organic medium such as fats, waxes, oil, fuel, petrol and the like are contained in esterified form (i.e. are bound in esters), a saponification step can be performed before the inventive removal and solubilization is carried out. Such a saponification is preferably performed in a solvent mixture of water and at least a second solvent miscible with water. Further preferred carboxylic acids are perfluoro carboxylic acids such perfluoropropionic acid, perfluorooctanoic acid (PFOA), perfluorodecanoic acid, perfluorododecanoic acid, perfluorohexadecanoic acid as well as other perfluoro carboxylic acids and prophyrinic acid.
The present invention also refers to the solubilization, respectively the removal of aromatic carboxylic acids belonging to the above-mentioned target groups, such as benzoic acid, 4-aminobenzoic acid, anthranilic acid, benzilic acid, cinnamic acid, salicylic acid, phenylacetic acid, 4-methoxy-phenylacetic acid, gallic acid, phthalic acid, terephthalic acid, abietic acid, bicinchoninic acid, quinic acid, chorismic acid, clavulanic acid, fusaric acid, fusidic acid, uric acid, hippuric acid, ibotenic acid, indole-3-acetic acid, mandelic acid, styphnic acid, usnic acid, abscisic acid, tropic acid, benzoquinonetetracarboxylic acid, boswellic acid, caffeic acid, carminic acid, chenodeoxycholic acid, coumaric acid, cromoglicic acid, cynarine, meclofenamic acid, 2,4-dichlorophenoxyacetic acid, domoic acid, pipemidic acid, ferulic acid, 5-hydroxyferulic acid, isophthalic acid, mefenamic acid, meta-chloroperoxybenzoic acid, peroxybenzoic acid, protocatechuic acid, nalidixic acid, sinapic acid, sucrononic acid.
Especially preferred is the removal and solubilization of carboxylic acids from blood which lead to various diseases caused and/or associated by an increased and/or unhealthy level of such carboxylic acids and especially fatty acids.
The carboxylic acids are preferably lipophil and preferably have a partition coefficient between n-octanol and water (also known as log KOW or octanol-water-partition coefficient) of >2.0, preferably of >3.0 and more preferably of >4.0. (For example: log KOW of acetic acid is −0.17, of butyric acid is 0.79, of octanoic acid is 3.05 and of decanoic acid is 4.09).
It is also preferred if the carboxylic acids which shall be removed have an pKs value >4.85, preferably >4.87. (for instance: acetic acid has pKs of 4.76, butyric acid of 4.82, pentanoic acid of 4.84 and octanoic acid of 4.89).
Thus the present invention provides a method for separating carboxylic acids which are not at all or not good soluable in water and which can be solubilized in water by means of the solubilizing compounds disclosed herein preferably in form of nano- or microemulsions. Once transferred into the aqueous phase, the fatty acids can be removed by various technics disclosed herein.
Thus the present invention relates to the use of a solubilizing compound for solubilizing carboxylic acids in an aqueous or organic medium, wherein said solubilizing compound contains at least one amidino group and/or at least one guanidino group and wherein the compound has a partition coefficient between n-octanol and water of KOW<6.30.
The term “solubilizing carboxylic acids in an aqueous or organic medium” should be understood as follows: the carboxylic acids which shall be solubilized are contained in an organic medium such as oils or fuel or in an aqueous medium such as blood or milk and are solubilized by the use of a solubilizing compound in the aqueous phase.
Thus it can also be stated that the present invention is directed to the use of a solubilizing compound for solubilizing carboxylic acids from an aqueous or organic medium in an aqueous phase, wherein said solubilizing compound contains at least one amidino group and/or at least one guanidino group and wherein the compound has a partition coefficient between n-octanol and water of KOW<6.30.
Moreover the present invention relates to the use of a solubilizing compound for solubilizing lipophilic carboxylic acids in an aqueous medium, wherein said solubilizing compound contains at least one amidino group and/or at least one guanidino group and wherein the compound has a partition coefficient between n-octanol and water of KOW<6.30.
In case where the carboxylic acids are contained in the aqueous phase such as blood, only very few amounts of free carboxylic acids are present in the blood, since these carboxylic acids and especially fatty acids are poorly water soluable. Most of the carboxylic acids which should be removed from the blood are bound to other compounds such as albumin and are no longer free carboxylic acids. However there is an equilibrium between the very small amount of free carboxylic acids in the blood and the otherwise bound or deposited carboxylic acids which are regarded as not free anymore. If now by means of the inventive method the free carboxylic acids are complexed by the solubilizing compound, these free carboxylic acids are removed from the equilibrium and albumin bound carboxylic acids are released into the blood which than can again be removed by the inventive method so that finally almost all carboxylic acids contained in the blood in a free or bound form can be removed. Especially dialysis is suitable for such a continuous process of removing the carboxylic acids and especially fatty acids from the blood.
The solubilizing compounds disclosed herein comprise at least one amidino group or at least one guanidino group or at least one amidino group and at least one guanidino group. If the amidino group is not substituted it can be represented by the following formula H2N—C(NH)—. But it is also possible that all three hydrogen atoms are replaced by substituents R, R′ and R″ as represented by the following general formula (R)(R′)N—C(NR″)—. It is preferred if two of the three hydrogen atoms are replaced by a substituent as represented by the following formula: (R′)NH—C(NR″)— or (R)(R′)N—C(NH)—. Thus amidino groups with at least one hydrogen are preferred. If the guanidino group is not substituted it can be represented by the following formula H2N—C(NH)—NH—. But it is also possible that all four hydrogen atoms are replaced by substituents R, R′, R″ and R′″ as represented by the following formula (R)(R′)N—C(NR″)—N(R″″)—. It is preferred if three of the four hydrogen atoms are replaced by a substituent as represented by the following formula: (R′)NH—C(NR″)—N(R″″)— or (R)(R′)N—C(NH)—N(R″″)— or (R)(R′)N—C(NR″)—NH—. Thus guanidino groups with at least one hydrogen and preferably with two hydrogens are preferred.
The solubilizing compound comprises or contains at least one amidino group and/or at least one guanidino group, while guanidino groups are preferred. Moreover the solubilizing compound comprises or contains preferably not more than 15 carbon atoms, more preferably not more than 14, more preferably not more than 13, more preferably not more than 12, more preferably not more than 11, more preferably not more than 10, more preferably not more than 9, and more preferably not more than 8 carbon atoms and most preferably the solubilizing compound is an arginine derivative. In case of polymeric or oligomeric solubilizing compounds it is preferred that per amidino moiety or per guanidino moiety not more than 10 carbon atoms and more preferably not more than 8 carbon atoms are present.
Furthermore the solubilizing compound is hydrophil and may preferably contain one or more of the following substituents:
—NH2, —OH, —PO3H2, —PO3H−, —PO32−, —OPO3H2, —OPO3H, —OPO32, —COOH, —COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO3, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH.
Also preferred are solubilizing compounds which are derivatives of arginine or which are dipeptides or tripeptides or polypeptides containing the amino acid arginine or a derivative of arginine.
It is also possible that the amidino group or the guanidino group is part of a heterocyclic ring system like in imidazole, histidine, clothianidin or 4-(4,5-dihydro-1H-imidazol-2-ylamino)-butyric acid.
The solubilizing compounds are hydrophil and have a partition coefficient between n-octanol and water (also known as KOW or octanol-water-partition coefficient) of KOW<6.30 (log KOW<0.80), preferably KOW<1.80 (log KOW<0.26), more preferably KOW<0.63 (log KOW<−0.20) and most preferably KOW<0.40 (log KOW<−0.40).
Preferred solubilizing compounds are:
L-2-amino-3-guanidinopropionic acid, L-arginine, L-NIL, H-homoarg-OH, histidine, Nω-nitro-L-arginine, N-ω-hydroxy-L-norarginine, D-arginine methyl ester, nomega-monomethyl-L-arginine, NG,NG-dimethylarginine, D-(+)-octopine, argininosuccinic acid, L-canavanine free base, creatine, guanidinoacetic acid, 3-guanidinopropionic acid, 4-guanidinobutyric acid, 4-(4,5-dihydro-1H-imidazol-2-ylamino)-butyricacid, (S)-(−)-2-guanidinoglutaric acid, 6-guanidinohexanoic acid, guanidino, sulfaguanidine, agmatinsulfate, 4-guanidinobenzoicacid, 1,3-di-o-tolyl-guanidine, clothianidine, L-ornitin, N-guanylurea, cimetidine, 1-(o-tolyl)biguanide, chlorhexidine, 1,1-dimethylbiguanide, proguanil, polyhexanide, poly-L-arginine (70.000-150.000 mw), diminazene, melanine, 4-(4,6-diamino-2,2-dimethyl-2H-[1,3,5]triazine-1-yl, imidazole, methylimidazole, Tyr-Arg (Kyotorphin), Arg-Gln, Gly-Arg, Arg-Phe, Arg-Glu, Lys-Arg acetate, His-Arg, Arg-Gly-Asp (RGD), Arg-Phe-Ala, Thr-Lys-Pro-Arg (Tuftsin), Gly-Gly-Tyr-Arg, Gly-His, argatroban, L-NMMA (L-NG-monomethyl-arginine), L-NAME (L-nitro-arginin-methylester), L-hydroxy-arginin-citrate, dimethylarginine (ADMA), D-homoarginine, noraginine, L-canavanin (2-amino-4-(guanidinooxy)-butyric acid), 4-guanidino-phenylalanine, 3-guanidino-phenylalanine, O-α-hippuryl-L-argininic acid, H-Arg-AMC (L-arginin-7-amido-4-methylcumarin), L-TAME (P-tosyl-L-arginin-methylester), diphenylacetyl-D-Arg-4-hydroxybenzylamid, agmatin (argamin; 1-amino-4-guanidinobutansulfate), L-arginin-ethylester, L-arginin-methylester, guanidine, guanidinacetate, guanidincarbonate, guanidinnitrate, guanidinthiocyanate, guanyl urea, guanyl urea phosphate, guanyl urea dinitramide, 2-guanidinoacetaldehyd-diethylacetale, dicyandiamide, 2-guanidinobenzimidazol, S-((2-guanidino-4-thiazolyl)methyl)-isothio urea, guanidinobutylaldehyde, 4-guanidinobenzoic acid, leonurin (4-guanidino-n-butylsyringate), ambazon ([4-(2-(diaminomethyliden)-hydrazinyl)phenyl]iminothio urea), amilorid (3,5-diamino-N-carbamimidoyl-6-chlorpyrazin-2-carbamide), aminoguanidine, amitrol (3-amino-1,2,4-triazole), nitroguanidine, argininosuccinate, barettin ((2S,5Z)-cyclo-[(6-brom-8-en-tryptophan)-arginine]), lysine, chlorhexidine (1,1′-hexamethylenbis[5-(4-chlorphenyl)-biguanide]), cimetidine (2-cyan-1-methyl-3-[2-(5-methylimidazol-4-ylmethylsulfanyl)-ethyl]-guanidine, clonidin (2-[(2,6-dichlorphenyl)imino]imidazolidine), clothianidin ((E)-1-(2-chlor-1,3-thiazol-5-ylmethyl)-3-methyl-2-nitroguanidine), 2,4-diaminopyridine, N,N′-di-o-tolylguanidine, guanethidine, kreatin, kreatinin, kyotorphin (L-tyrosyl-L-arginine), lugdunam, (N-(4-cyanophenyl)-N-(2,3-methylendioxybenzyl)guanidinacetic acid, metformin (1,1-dimethylbiguanid), octopin (Na-(1-carboxyethyl)arginine), polyhexanide (polyhexamethylenbiguanide (PHMB)), proguanil (1-(4-chlorphenyl)-5-isopropylbi-guanide), sulfaguanidine (4-amino-N-(diaminomethylen)benzensulfonamide), tetrazene (4-amidino-1-(nitrosaminoamidino)-1-tetrazena), L-arginine-4-methoxy-β-naphthylamide, L-arginine-β-naphthylamide, L-arginin-hydroxamate, L-arginine-p-nitroanilid, N-α-benzoyl-DL-arginine, Nω-Nitro-L-arginine, robenidin, (1,3-bis[(4-chlorobenzyliden)amino]-guanidine, 1-(2,2-diethoxyethyl)guanidine, 1-(P-tolyl)-guanidine nitrate.

The invention can be effectively used over a broad range of concentration ratios of the solubilizing compound and the fatty acids to be solubilized. Often the fatty acid content of a solution is not exactly known. Therefore the ratio of the solubilizing compound to be added has to be estimated. Inventive solubilization of carboxylic acids and especially fatty acids can be reached when the molar ratio of solubilizing compound to fatty acids (free and bounded) is in the range of 1:1000 to 1000:1. Preferred is a range of 1:100 to 100:1. More preferred is a range of 1:10 to 10:1. Further preferred is a range of 1:2 to 2:1. Most preferred is a ratio of 1:1 to 2:1. It is preferred that the solubilizing compound is used in a molar excess of 3% or 5% or 7% or 8% or 10% or 12% or 15% or 20% or 25% or 30% or 35% or 40% or 45% or 50% or 55% or 60% or 70% or 80% or 90% or 100% or 120% or 140% or 160% or 180% or 200% Moreover a molar ratio of fatty acid to solubilizing compound of 1:1 to 1:200 is preferred. More preferred is a molar ratio of fatty acid to solubilizing compound in the range of 1:1 to 1:100, more preferred of 1:1 to 1:50, still more preferred of 1:1 to 1:30, still more preferred of 1:1 to 1:25, still more preferred of 1:1 to 1:20, still more preferred of 1:1 to 1:15, still more preferred of 1:1 to 1:10, still more preferred of 1:1 to 1:9, still more preferred of 1:1 to 1:8, still more preferred of 1:1 to 1:7, still more preferred of 1:1 to 1:6, still more preferred of 1:1 to 1:5, still more preferred of 1:1 to 1:4, still more preferred of 1:1 to 1:3, still more preferred of 1:1 to 1:2, still more preferred of 1:1 to 1:1.8, still more preferred of 1:1 to 1:1.6, still more preferred of 1:1 to 1:1.5, still more preferred of 1:1 to 1:1.4, also preferred of 1:1 to 1:1.3, also preferred of 1:1 to 1:1.2, also preferred of 1:1 to 1:1.1, also preferred of 1:1 to 1:1.05, also preferred of 1:1.2 to 1:2.8, also preferred of 1:1.4 to 1:2.6, also preferred of 1:1.6 to 1:2.4, also preferred of 1:1.8 to 1:2.2, more preferred of 1:1.9 to 1:2.1 and most preferred is a molar ratio of fatty acid to solubilizing compound in the range of 1.0:2.0. These molar ratios are preferably for solubilizing compounds with one amidino group or one guanidino group. If the solubilizing compound contains two amidino groups or two guanidino groups or one amidino group and one guanidino group only half of the amount of the solubilizing compound is preferably used. Thus in such a case a molar ratio of fatty acid to solubilizing compound of 1:0.5 to 1:25, preferably 1:0.6 to 1:1.4, also preferred of 1:0.7 to 1:1.3, also preferred of 1:0.8 to 1:1.2, also preferred of 1:0.9 to 1:1.1, more preferred of 1:0.95 to 1:1.05 and most preferred is a molar ratio of fatty acid to solubilizing compound in the range of 1.0:1.0.
The solubilization is preferably carried out at a pH value >7.0 and more preferably within a pH range of 7.0 to 9.0. However depending on the medium from which the carboxylic acids should be separated, pH values up to 14 can be used, while a pH range between 7.0 and 8.0 is preferably used if the carboxylic acids should be removed from blood. However, if not complete solubilization is obtained, more solubilizing compound might be added or the pH value might be increased or the aqueous layer might be separated and the extraction process is repeated or a combination of these three possibilities is used.
Some of the solubilizing compounds of the present invention can be represented by the following general formula (I) and formula (II):

wherein
R′, R″, R′″ and R″″ represent independently of each other —H, —OH, —CH═CH2, —CH2—CH═CH2, —C(CH3)═CH2, —CH═CH—CH3, —C2H4—CH═CH2, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —C(CH3)3, —C5H11, —CH(CH3)—C3H7, —CH2—CH(CH3)—C2H5, —CH(CH3)—CH(CH3)2, —C(CH3)2—C2H5, —CH2—C(CH3)3, —CH(C2H5)2, —C2H4—CH(CH3)2, —C6H13, —C7H15, cyclo-C3H5, cyclo-C4H7, cyclo-C5H9, cyclo-C6H11, —PO3H2, —PO3H−, —PO32, —NO2, —C≡CH, —C≡C—CH3, —CH2—C≡CH, —C2H4—C≡CH, —CH2—C≡C—CH3, or R′ and R″ form together the residue —CH2—CH2—, —CO—CH2—, —CH2—CO—, —CH—CH—, —CO—CH—CH—, —CH—CH—CO—, —CO—CH2—CH2—, —CH2—CH2—CO—, —CH2—CO—CH2— or —CH2—CH2—CH2—
X represents —NH—, —NR″″—, —O—, —S— or —CH2— or a substituted carbon atom; and
L represents a hydrophilic substituent selected from the group comprising or consisting of
—NH2, —OH, —PO3H2, —PO3, —PO, —OPO3H2, —OPO3H−, —OPO32−, —COOH, −COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO3, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH, or

or
L represents a C1 to C8 linear or branched and saturated or unsaturated carbon chain with at least one substituent selected from the group comprising or consisting of
—NH2, —OH, —PO3H2, —PO3H, —PO32, —OPO3H2, —OPO3H, —OPO32, —COOH, —COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO3−, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH, or

or
L represents a benzene ring and preferably a para substituted benzene ring with at least one substituent selected from the group comprising or consisting of
—NH2, —OH, —PO3H2, —PO3H−, —PO32−, —OPO3H2, —OPO3H, —OPO32, —COOH, −COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO3, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH, or

However, such compounds are not preferred and can be excluded from the present application wherein X represents —O— or —S— and L represents —NH2, —OH, —OPO3H2, —OPO3H, —OPO32, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO3−, —SO2NH2, —CO—COOH, —O—CO—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH,

Also excluded are compounds wherein X represents —NH— or —NR″″— and L represents —OPO3H2, —OPO3H, —OPO32, —NH—CO—NH2, —CO—COOH, —O—CO—NH2, —NH—C(NH)—NH2, —NH—CS—NH2 or —NH—COOH.
The residue L may be further substituted by substituents as defined as R1 to R13. The residue L consists preferably of 1 to 10 carbon atoms, more preferably of 1 to 6 carbon atoms and most preferably of 2 to 4 carbon atoms. Carbon atoms of any substituents such as —COOH present on the residue L are included in the aforementioned carbon atom number. Thus the residue L contains a linear or branched carbon atom chain or a phenyl ring which might be substituted with one or more saturated or unsaturated and linear or branched alkyl substituents and/or substituents defined as R1 to R13.
It is preferred that the carbon chain of L is in the range of C1 to C7, more preferred in the range of C1 to C5 and most preferred in the range of C1 to C5.
Compounds of general formula (I) or (II) which can be used for solubilizing fatty acids in an aqueous medium or in water are represented by the following formula (I) or (II):

wherein
R′, R″, R′″ and R″″ represent independently of each other —H, —OH, —CH═CH2, —CH2—CH═CH2, —C(CH3)═CH2, —CH═CH—CH3, —C2H4—CH═CH2, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —C(CH3)3, —C5H11, —CH(CH3)—C3H7, —CH2—CH(CH3)—C2H5, —CH(CH3)—CH(CH3)2, —C(CH3)2—C2H5, —CH2—C(CH3)3, —CH(C2H5)2, —C2H4—CH(CH3)2, —C6H13, —C7H15, cyclo-C3H5, cyclo-C4H7, cyclo-C5H9, cyclo-C6H11, —PO3H2, —PO3H−, —PO32−, —NO2, —C≡CH, —C═C—CH3, —CH2—C≡CH, —C2H4—C≡CH, —CH2—C≡C—CH3, or R′ and R″ form together the residue —CH2—CH2—, —CH═CH— or —CH2—CH2—CH2—X represents —NH—, —NR″″, O, S or —CH2— or a substituted carbon atom; and
L represents —CR1R2R3, —CR4R5—CR1R2R3, —CR6R7—CR4R5—CR1R2R3, —CR6R9—CR6R7—CR4R5—CR1R2R3, —CR8R9—CR6R7—CR4R5—CR1R2R3, —CR12R13—CR19R11—CR8R9—CR6R7—CR4R5—CR1R2R3;

R*, R#, R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13 represent independently of each other the following substituents:
—NH2, —OH, —PO3H2, —PO3H, —PO32−, —OPO3H2, —OPO3H, —OPO32−, —COOH, —COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H5)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH, —H, —OCH3, —OC2H5, —OC3H7, -β-cyclo-C3H5, —OCH(CH3)2, —P(O)(OCH3)2, —Si(CH3)2(C(CH3)3), —OC(CH3)3, —OC4H9, —OPh, —OCH2-Ph, —OCPh3, —SH, —SCH3, —SC2H5, —SC3H7, —S-cyclo-C3H5, —SCH(CH3)2, —SC(CH3)3, —NO2, —F, —Cl, —Br, —I, —P(O)(OC2H5)2, —P(O)(OCH(CH3)2)2, —C(OH)[P(O)(OH)2]2, —Si(C2H5)3, —Si(CH3)3, —N3, —CN, —OCN, —NCO, —SCN, —NCS, —CHO, —COCH3, —COC2H5, —COC3H7, —CO-cyclo-C3H5, —COCH(CH3)2, —COC(CH3)3, —COCN, —COOCH3, —COOC2H5, —COOC3H7, —COO-cyclo-C3H5, —COOCH(CH3)2, —COOC(CH3)3, —OOC—CH3, —OOC—C2H5, —OOC—C3H7, —OOC-cyclo-C3H5, —OOC—CH(CH3)2, —OOC—C(CH3)3, —CONHCH3, —CONHC2H5, —CONHC3H7, —CONH-cyclo-C3H5, —CONH[CH(CH3)2], —CONH[C(CH3)3], —CON(CH3)2, —CON(C2H5)2, —CON(C3H7)2, —CON(cyclo-C3H5)2, —CON[CH(CH3)2]2, —CON[C(CH3)3]2, —NHCOCH3, —NHCOC2H5, —NHCOC3H7, —NHCO-cyclo-C3H5, —NHCO—CH(CH3)2, —NHCO—C(CH3)3, —NHCO—OCH3, —NHCO—OC2H5, —NHCO—OC3H7, —NHCO-β-cyclo-C3H5, —NHCO—OCH(CH3)2, —NHCO—OC(CH3)3, —NH-cyclo-C3H5, —NHCH(CH3)2, —NHC(CH3)3, —N(CH3)2, —N(C2H5)2, —N(C3H7)2, —N(cyclo-C3H5)2, —N[CH(CH3)2]2, —N[C(CH3)3]2, —SOCH3, —SOC2H5, —SOC3H7, —SO-Cyclo-C3H5, —SOCH(CH3)2, —SOC(CH3)3, —SO2CH3, —SO2C2H5, —SO2C3H7, —SO2-cyclo-C3H5, —SO2CH(CH3)2, —SO2C(CH3)3, —SO3CH3, —SO3C2H5, —SO3C3H7, —SO3-cyclo-C3H5, —SO3CH(CH3)2, —SO3C(CH3)3, —SO2NH2, —OCF3, —OC2H5, —O—COOCH3, —O—COOC2H5, —O—COOC3H7, —O—COO-cyclo-C3H5, —O—COOCH(CH3)2, —O—COOC(CH3)3, —NH—CO—NHCH3, —NH—CO—NHC2H5, —NH—CS—N(C3H7)2, —NH—CO—NHC3H7, —NH—CO—N(C3H7)2, —NH—CO—NH[CH(CH3)2], —NH—CO—NH[C(CH3)3], —NH—CO—N(CH3)2, —NH—CO—N(C2H5)2, —NH—CO—NH-cyclo-C3H5, —NH—CO—N(cyclo-C3H5)2, —NH—CO—N[CH(CH3)2]2, —NH—CS—N(C2H5)2, —NH—CO—N[C(CH3)3]2, —NH—CS—NH2, —NH—CS—NHCH3, —NH—CS—N(CH3)2, —NH—CS—NHC2H5, —NH—CS—NHC3H7, —NH—CS—NH-cyclo-C3H5, —NH—CS—NH[CH(CH3)2], —NH—CS—NH[C(CH3)3], —NH—CS—N(cyclo-C3H5)2, —NH—CS—N[CH(CH3)2]2, —NH—CS—N[C(CH2)2]2, —NH—C(═NH)—NH2, —NH—C(═NH)—NHCH3, —NH—C(═NH)—NHC2H5, —NH—C(═NH)—NHC3H7, —O—CO—NH-cyclo-C3H5, —NH—C(═NH)—NH-cyclo-C3H5, —NH—C(═NH)—NH[CH(CH3)2], —O—CO—NH[CH(CH3)2], —NH—C(═NH)—NH[C(CH3)3], —NH—C(═NH)—N(CH3)2, —NH—C(═NH)—N(C2H5)2, —NH—C(═NH)—N(C3H7)2, —NH—C(═NH)—N(cyclo-C3H5)2, —O—CO—NHC3H7, —NH—C(═NH)—N[CH(CH3)2]2, —NH—C(═NH)—N[C(CH3)3]2, —O—CO—NHCH3, —O—CO—NHC2H5, —O—CO—NH[C(CH3)3], —O—CO—N(CH3)2, —O—CO—N(C2H5)2, —O—CO—N(C3H7)2, —O—CO—N(cyclo-C3H5)2, —O—CO—N[CH(CH3)2]2, —O—CO—N[C(CH3)3]2, —O—CO—OCH3, —O—CO—OC2H5, —O—CO—OC3H7, —O—OO—O-cyclo-C3H5, —O—CO—OCH(CH3)2, —O—CO—OC(CH3)3, —CH2F, —CHF2, —CF3, —CH2Cl, —CH2Br, —CH2I, —CH2—CH2F, —CH2—CHF2, —CH2—CF3, —CH2—CH2Cl, —CH2—CH2Br, —CH2—CH2I, cyclo-C3H5, cyclo-C4H7, cyclo-C5H9, cyclo-C6H11, cyclo-C7H13, cyclo-C5H15, -Ph, —CH2-Ph, —CPh3, —CH3, —C2H5, —C3H7, —CH(CH3)2, —C4H9, —CH2—CH(CH3)2, —CH(CH3)—C2H5, —C(CH3)3, —C5H11, —CH(CH3)—C3H7, —CH2—CH(CH3)—C2H5, —CH(CH3)—CH(CH3)2, —C(CH3)2—C2H5, —CH2—C(CH3)3, —CH(C2H5)2, —C2H4—CH(CH3)2, —C6H13, —C7H15, —C8H7, —C3H5—CH(CH3)2, —C2H4—CH(CH3)—C2H5, —CH(CH3)—C4H9, —CH2—CH(CH3)—C3H7, —CH(CH3)—CH2—CH(CH3)2, —CH(CH3)—CH(CH3)—C2H5, —CH2—CH(CH3)—CH(CH3)2, —CH2—C(CH3)2—C2H5, —C(CH3)2—C3H7, —C(CH3)2—CH(CH3)2, —C2H4—C(CH3)3, —CH(CH3)—C(CH3)3, CH═CH2, —CH2—CH═CH2, —C(CH3)═CH2, —CH═CH—CH3, —C2H4—CH═CH2, —CH2—CH═CH—CH3, —CH═CH—C2H5, —CH2—C(CH3)═CH2, —CH(CH3)—CH═CH, —CH═C(CH3)2, —C(CH3)═CH—CH3, —CH═CH—CH═CH2, —C3H6—CH═CH2, —C2H4—CH═CH—CH3, —CH2—CH═CH—C2H5, —CH═CH—C3H7, —CH2—CH═CH—CH═CH2, —CH═CH—CH═CH—CH3, —CH═CH—CH2—CH═CH2, —C(CH3)═CH—CH═CH2, —CH═C(CH3)—CH═CH2, —CH═CH—C(CH3)═CH2, —C2H4—C(CH3)═CH2, —CH2—CH(CH3)—CH═CH2, —CH(CH3)—CH2—CH═CH2, —CH2—CH═C(CH3)2, —CH2—C(CH3)═CH—CH3, —CH(CH3)—CH═CH—CH3, —CH═CH—CH(CH3)2, —CH═C(CH3)—C2H5, —C(CH3)═CH—C2H5, —C(CH3)═C(CH3)2, —C(CH3)2—CH═CH2, —CH(CH3)—C(CH3)═CH2, —C(CH3)═CH—CH═CH2, —CH═C(CH3)—CH═CH2, —CH═CH—C(CH3)═CH2, —C4H5—CH═CH2, —C3H6—CH═CH—CH3, —C2H4—CH═CH—C2H5, —CH2—CH═CH—C3H7, —C3H6—C(CH3)═CH2, —C2H4—CH(CH3)—CH═CH2, —CH2—CH(CH3)—CH2—CH═CH2, —C2H4—CH═C(CH3)2, —CH(CH3)—C2H4—CH═CH2, —C2H4—C(CH3)═CH—CH3, —CH2—CH(CH3)—CH═CH—CH3, —CH(CH3)—CH2—CH═CH—CH3, —CH2—CH═CH—CH(CH3)2, —CH2—CH═C(CH3)—C2H5, —CH2—C(CH3)═CH—C2H5, —CH(CH3)—CH═CH—C2H5, —CH═CH—CH2—CH(CH3)2, —CH═CH—CH(CH3)—C2H5, —CH═C(CH3)—C3H7, —C(CH3)═CH—C3H7, —CH2—CH(CH3)—C(CH3)═CH2, —C[C(CH3)3]═CH2, —CH(CH3)—CH2—C(CH3)═CH2, —CH(CH3)—CH(CH3)—CH═CH2, —CH═CH—C2H4—CH═CH2, —CH2—C(CH3)2—CH═CH2, —C(CH3)2—CH2—CH═CH2, —CH2—C(CH3)═C(CH3)2, —CH(CH3)—CH═C(CH3)2, —C(CH3)2—CH═CH—CH3, —CH═CH—CH2—CH═CH—CH3, —CH(CH3)—C(CH3)═CH—CH3, —CH═C(CH3)—CH(CH3)2, —C(CH3)═CH—CH(CH3)2, —C(CH3)═C(CH3)—C2H5, —CH═CH—C(CH3)3, —C(CH3)2—C(CH3)═CH2, —CH(C2H5)—C(CH3)═CH2, —C(CH3)(C2H5)—CH═CH2, —CH(CH3)—C(C2H5)═CH2, —CH2—C(C3H7)═CH2, —CH2—C(C2H5)═CH—CH3, —CH(C2H5)—CH═CH—CH3, —C(C4H9)═CH2, —C(C3H7)═CH—CH3, —C(C2H5)═CH—C2H5, —C(C2H5)═C(CH3)2, —C[CH(CH3)(C2H5)]═CH2, —C[CH2—CH(CH3)2]═CH2, —C2H4—CH═CH—CH═CH2, —CH2—CH═CH—CH2—CH═CH2, —C3H6—C≡C—CH3, —CH2—CH═CH—CH═CH—CH3, —CH═CH—CH═CH—C2H5, —CH2—CH═CH—C(CH3)═CH2, —CH2—CH═C(CH3)—CH═CH2, —CH2—C(CH3)═CH—CH═CH2, —CH(CH3)—CH2—C≡CH, —CH(CH3)—CH═CH—CH═CH2, —CH═CH—CH2—C(CH3)═CH2, —CH(CH3)—C≡C—CH3, —CH═CH—CH(CH3)—CH═CH2, —CH═C(CH3)—CH2—CH═CH2, —C2H4—CH(CH3)—C≡CH, —C(CH3)═CH—CH2—CH═CH2, —CH═CH—CH═C(CH3)2, —CH2—CH(CH3)—CH2—C≡CH, —CH═CH—C(CH3)═CH—CH3, —CH═C(CH3)—CH═CH—CH3, —CH2—CH(CH3)—C═CH, —C(CH3)═CH—CH═CH—CH3, —CH═C(CH3)—C(CH3)═CH2, —C(CH3)═CH—C(CH3)═CH2, —C(CH3)═C(CH3)—CH═CH2, —CH═CH—CH═CH—CH═CH2, —C≡CH, —C≡C—CH3, —CH2—C≡CH, —C2H4—C≡CH, —CH2—C═C—CH3, —C═C—C2H5, —C3H6—C≡CH, —C2H4—C═C—CH3, —CH2—C═C—C2H5, —C═C—C3H7, —CH(CH3)—C≡CH, —C4H5—C≡CH, —C2H4—C═C—C2H5, —CH2—C═C—C3H7, —C═C—C4H3, —C≡C—C(CH3)3, —CH(CH3)—C2H4—C≡CH, —CH2—CH(CH3)—C═C—CH3, —CH(CH3)—CH2—C═C—CH3, —CH(CH3)—C≡C—C2H5, —CH2—C═C—CH(CH3)2, C≡C—CH(CH3)—C2H5, —C≡C—CH2—CH(CH3)2, —CH(C2H5)—C≡C—CH3, —C(CH3)2—C≡C—CH3, —CH(C2H5)—CH2—C≡CH, —CH2—CH(C2H5)—C≡CH, —C(CH3)2—CH2—C≡CH, —CH2—C(CH3)2—C≡CH, —CH(CH3)—CH(CH3)—C══CH, —CH(C3H7)—C≡CH, —C(CH3)(C2H5)—C≡CH, —CH2—CH(C≡CH)2, —C≡C—C≡CH, —CH2—C≡C—C≡CH, —C≡C—C≡C—CH3, —CH(C≡CH)2, —C2H4—C≡C—C≡CH, —CH2—C≡C—CH2—C≡CH, —C≡C—C2H4—C≡CH, —CH2—C≡C—C≡C—CH3, —C≡C—CH2—C≡C—CH3, —C═C—C≡C—C2H5, —C(C≡CH)2—CH3, —C═C—CH(CH3)—C≡CH, —CH(CH3)—C≡C—C≡CH, —CH(C≡CH)—CH2—C≡CH, —CH(C≡CH)—C═C—CH3, —CH═CH-Ph, —NH—CO—CH2—COOH, —NH—CO—C2H4—COOH, —NH—CO—CH2—NH2, —NH—CO—C2H4—NH2, —NH—CH(COOH)—CH2—COOH, —NH—CH2—COOH, —NH—C2H4—COOH, —NH—CH(COOH)—C2H4—COOH, —NH—CH(CH3)—COOH;
wherein preferably at least one of the substituents R*, R#, R1, R2, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13 is selected from the following substituents:
—NH2, —OH, —PO3H2, —PO3H, —PO32−, —OPO3H2, —OPO3H, —OPO32−, —COOH, —COO−, —CO—NH2, —NH3+, —NH—CO—NH2, —N(CH3)3+, —N(C2H5)3+, —N(C3H7)3+, —NH(CH3)2+, —NH(C2H8)2+, —NH(C3H7)2+, —NHCH3, —NHC2H5, —NHC3H7, —NH2CH3+, —NH2C2H5+, —NH2C3H7+, —SO3H, —SO2NH2, —CO—COOH, —O—CO—NH2, —C(NH)—NH2, —NH—C(NH)—NH2, —NH—CS—NH2, —NH—COOH
Preferred are also compounds of the general formula (III) as shown below:

wherein the residues X and L have the meanings as disclosed herein.
Preferably the compounds of general formula (I), (II) and (III) have a partition coefficient between n-octanol and water (also known as KOW or octanol-water-partition coefficient) of KOW<6.30 (log KOW<0.80), preferably KOW<1.80 (log KOW<0.26), more preferably KOW<0.63 (log KOW<−0.20) and most preferably KOW<0.40 (log KOW<−0.40).
Moreover the compounds of general formula (I), (II) and (III) have the same preferred carbon atom number as disclosed above, the same preferred pH range for the solubilization reaction, the same preferred molar ratio of carboxylic acid to solubilization compound and the same preferred reaction conditions as disclosed above for the solubilization compounds in general.
The partition coefficient is a ratio of concentrations of the un-ionized compound between the two solutions. To measure the partition coefficient of ionizable solutes, the pH of the aqueous phase is adjusted such that the predominant form of the compound is un-ionized. The logarithm of the ratio of the concentrations of the un-ionized solute in the solvents is called log P:
      log    ⁢                  ⁢          P              oct        /        wat              =      log    ⁡          (                                    [            solute            ]                    octanol                                      [            solute            ]                    water                      un            -            ionized                              )      
The distribution coefficient is the ratio of the sum of the concentrations of all forms of the compound (ionized plus un-ionized) in each of the two phases. For measurements of the distribution coefficient, the pH of the aqueous phase is buffered to a specific value such that the pH is not significantly perturbed by the introduction of the compound. The logarithm of the ratio of the sum of concentrations of the various forms of the solute in one solvent, to the sum of the concentrations of its forms in the other solvent is called log D:
      log    ⁢                  ⁢          D              oct        /        wat              =      log    ⁡          (                                    [            solute            ]                    octanol                                                    [              solute              ]                        water            ionized                    +                                    [              solute              ]                        water            neutral                              )      
In addition, log D is pH dependent, hence the pH must be specified at which log D was measured. Of particular interest is the log D at pH=7.4 (the physiological pH of blood serum). For un-ionizable compounds, log P=log D at any pH.